Vitamin E is an essential component of mammalian diets. Epidemiological evidence indicates that Vitamin E supplementation results in decreased risk for cardiovascular disease and cancer, aids in immune function, and generally prevents or slows a number of degenerative disease processes in humans (Traber and Sies, Annu. Rev. Nutr. 16:321-347, 1996). Vitamin E functions in stabilizing the lipid bilayer of biological membranes (Skrypin and Kagan, Biochim. Biophys. Acta 815:209 1995; Kagan, N.Y. Acad. Sci. p 121, 1989; Gomez-Fernandez et al., Ann. N.Y. Acad. Sci. p 109, 1989), reducing polyunsaturated fatty acid (PUFA) free radicals generated by lipid oxidation (Fukuzawa et al., Lipids 17: 511-513, 1982), and quenching singlet oxygen species (Fryer, Plant Cell Environ. 15(4):381-392, 1992).
Vitamin E, or α-tocopherol, belongs to a class of lipid-soluble antioxidants that includes α, β, γ, and δ-tocopherols and α, β, γ, and δ-tocotrienols. Although α, β, γ, and δ-tocopherols and α, β, γ, and δ-tocotrienols are sometimes referred to collectively as “Vitamin E” in the popular press, Vitamin E is properly defined chemically solely as α-tocopherol. Of the various tocopherols present in foodstuff, α-tocopherol is the most significant for human health both because it is the most bioactive of the tocopherols and also because it is the tocopherol most readily absorbed and retained by the body (Traber and Sies, Annu. Rev. Nutr. 16:321-347, 1996). The in vivo antioxidant activity of α-tocopherol is higher than the antioxidant activities of β, γ, and δ-tocopherol (Kamal-Eldin and Appelqzvist Lipids 31:671-701, 1996).
Only plants and certain other photosynthetic organisms, including cyanobacteria, synthesize tocopherols. Therefore, dietary tocopherols are obtained almost exclusively from plants. Plant tissues vary considerably in total tocopherol content and tocopherol composition. The predominant tocopherol in green, photosynthetic plant tissues often is α-tocopherol. Leaf tissue can contain from 10-50 μg total tocopherols/gram fresh weight.
Non-green plant tissues and organs exhibit a wider range of both total tocopherol levels and tocopherol compositions. In general, most of the major food staple corps (e.g., rice, corn, wheat, potato) produce low to extremely low levels of total tocopherols, of which only a small percentage is α-tocopherol (Hess, Vitamin E, α-tocopherol, In Antioxidants in Higher Plants, R. Alscher and J. Hess, Eds. 1993, CRC Press, Boca Raton. pp 111-134). Oil seed crops generally contain much higher levels of total tocopherols; however, α-tocopherol is present only as a minor component and β, γ, and δ-tocopherols and tocotrienols predominate (Taylor and Barnes, Chemy Ind., Oct.:722-726, 1981).
Daily dietary intake of 15-30 mg of vitamin E is recommended to obtain optimal plasma α-tocopherol levels. It is quite difficult to achieve this level of vitamin E intake from the average American diet. For example, one could obtain the recommended daily dose of Vitamin E by daily consumption of over 750 grams of spinach leaves (in which α-tocopherol comprises 60% of total tocopherols) or 200-400 grams of soybean oil.
One alternative to relying on diet alone to obtain the recommended levels of vitamin E is to take a vitamin E supplement. However, most vitamin E supplements are synthetic vitamin E having six stereoisomers, whereas natural vitamin E vitamin is a single isomer. Furthermore, supplements tend to be relatively expensive, and the general population is disinclined to take vitamin supplements on a regular basis.
Although tocopherol function in plants has been less extensively studied than tocopherol function in mammalian systems, it is likely that the analogous functions performed by tocopherols in animals also occur in plants. In general, plant tocopherol levels have been found to increase with increases in various stresses, especially oxidative stress. Increased α-tocopherol levels in crops are associated with enhanced stability and extended shelf life of fresh and processed plant products (Peterson, Cereal-Chem 72(1):21-24, 1995; Ball, Fat-soluble vitamin assays in food analysis. A comprehensive review. London: Elsevier Science Publishers LTD, 1988).
Vitamin E supplementation of swine, beef, and poultry feeds has been shown to significantly increase meat quality and extend the shelf life of post-processed meat products by retarding post-processing lipid oxidation, which contributes to the formation of undesirable flavor components (Ball, supra 1988; Sante and Lacourt, J. Sci. Food Agric. 65(4):503-507, 1994; Buckley et al., J. of Animal Science 73:3122-3130, 1995).
What would be useful for the art is a method to increase the ratio of α-tocopherol to γ-tocopherol in seeds, oils, and leaves from crop and forage plants, or a method for producing natural vitamin E in nonphotosynthetic bacteria or fungi using a large scale fermentation process. Increasing α-tocopherol levels in crop plants would increase the amount of α-tocopherol obtained in the human diet, and would enhance the stability and shelf life of plants and plant products. The meat industry would benefit from the development of forage plants having increased levels of vitamin E.